Neonatal screening for isovaleric aciduria: Reducing the increasingly high false‐positive rate in Germany

Abstract

Newborn screening (NBS) for isovaleric acidemia (IVA) is performed by flow injection tandem mass spectrometry quantifying C5 carnitines (C5). Isovalerylcarnitine, however, is isomeric with pivaloylcarnitine which can be present in blood due to maternal use of pivaloylester-containing antibiotics, available in Germany since late 2016. During a 36-month period (January 19-December 21), all newborns screened in Hamburg with a C5 above cutoff (NeoGram ® : 0.50 μmol/L or Neobase ® 2: 0.45 μmol/L) were included in the study. As a second-tier test, a simple ultra performance liquid chromatography-tandem mass spectrometry (UPLC-MS/MS) method was developed to differentiate the C5 isomers pivaloyl-, 2-methylbutyryl-, isovaleryl-, and valerylcarnitine. Out of 156 772 newborns tested, one turned out to have genetically proven IVA while 99 were false positive (C5: 0.5-8.2 μmol/L) due to the presence of pivaloylcarnitine. These cases have increased year by year and show local clusters. Retrospective analysis of another 39 cases from 287 206 neonates tested at the NBS center in Heidelberg with C5 elevation (0.9-10.6 μmol/L) but clinical and biochemical exclusion of IVA yielded evidence of pivaloylcarnitine in all cases. Inclusion of a second-tier test into NBS significantly reduces the high and increasing false-positive rate of IVA screening. This avoids further diagnostic steps, prevents unnecessary stress and anxiety of parents in a remarkably high number of cases.If Hamburg data of 2021 are extrapolated to all of Germany, one can assume around 800 (1‰) false-positive cases in comparison to an average of two classic IVA cases per year. Unless licensing of pivaloylester-containing drugs for use during pregnancy is reconsidered, a second-tier test for C5 determination is indispensable.

Isovaleric aciduria identified by newborn screening: Strategies to predict disease severity and stratify treatment

“…An essential aim of any NBS program must be to prevent avoidable harm to patients and parents. 26 NBS for IVA is further hampered by the rising number of false positive screening results due to the increasing use of pivmecillinam-containing antibiotics in pregnant women, 9,28 which also requires further biochemical diagnostics to be excluded until possible upcoming biochemical second-tier 29 or computational approaches in pattern recognition as digital tier [30][31][32][33] tests are widely implemented.…”

“…This algorithm also includes the use of a second-tier test to discriminate between pivaloylcarnitine and C5 29 or a digital tier test. 31,32 Although, it is important to note that these additional tests should only be applied by NBS laboratories with established infrastructure to avoid any delay in reporting of the NBS results, they could be an important support to facilitate NBS for IVA and reduce harm to newborns with false positive results and their families.…”

Machine Learning Methods Improve Specificity in Newborn Screening for Isovaleric Aciduria

“…NBS for IVA in Germany is hampered by the identification of an attenuated disease variant in about of all newborns with confirmed IVA [ 4 ], for which the benefit of NBS is still unclear, and an increasing number of false positive screening results due to the use of pivmecillinam contained in the medication. [ 8 ]. This study examined the application of data analysis and ML methods as a potential digital-tier on NBS profiles to improve the specificity of NBS for IVA and support the classification of disease severity.…”

“…The application of ANOVA on the data sets confirmed biological knowledge such as highlighting C5, the known primary variable of the common NBS for IVA [ 4 , 5 , 9 ] as a significant feature for the full NBS data set, Table 2 . Moreover, on both data sets, ANOVA identified birth year as a significant feature, Table 2 , for all evaluations, which is explained by the increasing number of false positives in recent years, Supplementary Figure S1 , due to the use of pivmecillinam as an antibiotic in pregnant women since its authorization in Germany in March 2016 [ 8 , 33 ]. Other parameters were up to now not described to be altered in newborns with IVA nor are easy to explain regarding the metabolic pathways [ 4 , 5 , 9 ].…”

“…A multi-center long-term observational study showed individuals with mild IVA, of the screened with IVA in Germany, to be at risk for over-treatment [ 4 ]. The second struggle of NBS for IVA is an increasing number of false positives due to the increasing use of pivmecillinam, an antibiotic, used in urinary tract infections in pregnant women [ 8 ]. The antibiotic’s metabolite pivaloylcarnitine is isobaric to isovalerylcarnitine (C5), the primary biomarker in NBS for IVA [ 9 ].…”